The long term objective of this proposal is to elucidate the negative inotropic mechanism(s) by which the widely used vapor anesthetic agents (VAAs) halothane, enflurane and isoflurane, as well as the newer vapor agents sevoflurane and desflurane, depress cardiac and skeletal muscle contractility. More specifically, intracellular Ca2+ will be monitored in multicellular myocyte preparations with a battery of fluorescent Ca2+- reporter dyes and aequorin, the content of the sarcoplasmic reticulum (SR) will be assayed using the rapid cooling contracture technique, sarcolemmal influx of Ca2+ through L-type channels will be determined following electrical stimulation in the presence of ryanodine and commonly used indicators of contractility (e.g., force generation, speed of shortening ) will be recorded. The Specific Aims are: (1) to study the influence of the VAAs on these indicators of Ca2+ movement, SR content and contractility. This will provide new information about the specific mechanisms that produce the negative inotropic effects of the VAAs in the heart. (2) To use the same indicators of intracellular Ca2+ movement, SR content and contractility to clarify the mechanism of actio of various inotropic interventions in the heart, such as changes in extracellular [Ca2+] or changes in rate or pattern of stimulation, in order to compare similarities and differences with respect to VAA actions. (3) To establish the effects of the VAAs on excitation- contraction coupling in vertebrate skeletal muscle fibers and thus delineate similarities and differences with respect to those found in cardiac muscle. This last Specific Aim will also yield new information because the mechanism of excitation-contraction coupling in the two muscle types differs radically in that external Ca2+ is mandatory in heart while in skeletal muscle only a sarcolemmal voltage change is required. A key feature of the Research Design is the use of intact muscle where the intracellular milieu is most nearly physiological. The Methodology exploits many techniques that are already well developed in our laboratory. The major new technical advances, which we have utilized to obtain preliminary results, involve the use of intracellular fluorescent Ca2+ indicator dyes: Fura-2, Fluo-3, Mag-Fura-2 content. The unique feature of our proposal which adds much to its health relatedness is the intention to study the basis for the negative inotropic effect of the VAAs on contractility in both heart and skeletal muscle.